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Page 1: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase
Page 2: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

End of Lecture 1

Page 3: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

DiffractionScattering – a system of two electrons:

The two waves have equal amplitudes (because e1 and e2 are identical), but a phase difference

of 2r.S

|S|=|s-s0|=2sin/

(e1)Total scatter = (e1) + (e2)

= 1 + exp (2ir.S)

2r.S

(e2)

e1

e2

s0

s

r

Page 4: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

DiffractionScattering by an atom:

Atomic scattering factor

f(S) = r {exp (2ir.S) + exp (2i-r.S)} d3r

= r cos (2r.S) d3r

The atomic scattering factor is independent of the direction of S, but does depend on the length of S:

|S|=2sin/

Page 5: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Fibre Diffraction from Insect Flight Muscle - 1967

Page 6: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

The Beginning of Molecular Biology.

Francis Crick and James D Watson

Page 7: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

We wish to suggest a structure for the salt of deoxyribose nucleic acid (D.N.A.). This structure has novel features which are of considerable biological interest.

A structure for nucleic acid has already been proposed by Pauling and Corey (1). They kindly made their manuscript available to us in advance of publication. Their model consists of three intertwined chains, with the phosphates near the fibre axis, and the bases on the outside. In our opinion, this structure is unsatisfactory for two reasons: (1) We believe that the material which gives the X-ray diagrams is the salt, not the free acid. Without the acidic hydrogen atoms it is not clear what forces would hold the structure together, especially as the negatively charged phosphates near the axis will repel each other. (2) Some of the van der Waals distances appear to be too small.

Another three-chain structure has also been suggested by Fraser (in the press). In his model the phosphates are on the outside and the bases on the inside, linked together by hydrogen bonds. This structure as described is rather ill-defined, and for this reason we shall not comment on it.

We wish to put forward a radically different structure for the salt of deoxyribose nucleic acid. This structure has two helical chains each coiled round the same axis (see diagram). We have made the usual chemical assumptions, namely, that each chain consists of phosphate diester groups joining ß-D-deoxyribofuranose residues with 3',5' linkages. The two chains (but not their bases) are related by a dyad perpendicular to the fibre axis. Both chains follow right- handed helices, but owing to the dyad the sequences of the atoms in the two chains run in opposite directions. Each chain loosely resembles Furberg's model No. 1; that is, the bases are on the inside of the helix and the phosphates on the outside. The configuration of the sugar and the atoms near it is close to Furberg's 'standard configuration', the sugar being roughly perpendicular to the attached base. There is a residue on each every 3.4 A. in the z-direction. We have assumed an angle of 36° between adjacent residues in the same chain, so that the structure repeats after 10 residues on each chain, that is, after 34 A. The distance of a phosphorus atom from the fibre axis is 10 A. As the phosphates are on the outside, cations have easyaccess to them.

Page 8: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

The model of Pauling and Corey• consists of three intertwined chains, with phosphates

near the fibre axis, and the bases on the outside

• For Watson and Crick unsatisfactory because of:

1. Salt gives the X-ray diagram and not the free acid

2. It is not clear what forces hold the structure together, especially as the negativ charged phosphates near the axis will repel each other

3. Some of the van-der-Waals distance appear to small

Page 9: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

The structure is an open one, and its water content is rather high. At lower water contents we would expect the bases to tilt so that the structure could become more compact.

The novel feature of the structure is the manner in which the two chains are held together by the purine and pyrimidine bases. The planes of the bases are perpendicular to the fibre axis. The are joined together in pairs, a single base from the other chain, so that the two lie side by side with identical z-co-ordinates. One of the pair must be a purine and the other a pyrimidine for bonding to occur. The hydrogen bonds are made as follows : purine position 1 to pyrimidine position 1 ; purine position 6 to pyrimidine position 6.If it is assumed that the bases only occur in the structure in the most plausible tautomeric forms (that is, with the keto rather than the enol configurations) it is found that only specific pairs of bases can bond together. These pairs are : adenine (purine) with thymine (pyrimidine), and guanine (purine) with cytosine (pyrimidine).

In other words, if an adenine forms one member of a pair, on either chain, then on these assumptions the other member must be thymine ; similarly for guanine and cytosine. The sequence of bases on a single chain does not appear to be restricted in any way. However, if only specific pairs of bases can be formed, it follows that if the sequence of bases on one chain is given, then the sequence on the other chain is automatically determined .

Page 10: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

It has been found experimentally (3,4) that the ratio of the amounts of adenine to thymine, and the ration of guanine to cytosine, are always very close to unity for deoxyribose nucleic acid.

It is probably impossible to build this structure with a ribose sugar in place of the deoxyribose, as the extra oxygen atom wouldmake too close a van der Waals contact. The previously published X-ray data (5,6) on deoxyribose nucleic acid are insufficientfor a rigorous test of our structure. So far as we can tell, it is roughly compatible with the experimental data, but it must beregarded as unproved until it has been checked against more exact results. Some of these are given in the followingcommunications. We were not aware of the details of the results presented there when we devised our structure, which restsmainly though not entirely on published experimental data and stereochemical arguments.

It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanismfor the genetic material.

Full details of the structure, including the conditions assumed in building it, together with a set of co-ordinates for the atoms,will be published elsewhere.

Page 11: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

The model of Watson and Crick

• two helical chains each coiled round the same axis

• Right handed helices

• Residue on each chain every 3,4 Å in z direction

• Angle of 36° between the residues of the same chain

• Structure repeats after 10 residues, after 34 Å

• Bases are joined in pairs via hydrogen bond

• Pairs: adenine (purine) – thymine (pyrimidin), guanine (purine) – cytosine (pyrimidin)

Page 12: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

The model of Watson and Crick

D.N.A.:

BASE – SUGAR

PHOSPHATE

BASE – SUGAR

PHOSPHATE

BASE – SUGAR

PHOSPHATE

BASE – SUGAR

PHOSPHATE

Page 13: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

The model of Watson and Crick

Page 14: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase
Page 15: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

The model of Watson and Crick

Page 16: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Crystallography

Page 17: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

James Batcheller Sumner (1887-1955)Nobel prize in Chemistry 1946 for his discovery that enzymes can be crystallized

John Howard Northrop (1891-1987), Wendell Meredith Stanley (1904-1971) Nobel prize in Chemistry 1946 for their preparation of enzymes and virus proteins in a pure form

Page 18: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

John Desmond Bernal (1901-1971)Dorothy Crowfoot Hodgkin (1910-1994)

Page 19: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

X-ray crystallography - first purify and crystallize

Page 20: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Crystallography - Crystals

Page 21: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Crystallography

The main technique behind:

X-ray source: X-ray detector

Synchrotron Single photon counter

Storage ring photographic film

Rotating anode tube image plates

Sealed x-ray tube area detectors

Page 22: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase
Page 23: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase
Page 24: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Crystallography - Crystals

Unit cell:

a

b

c

a

b

c

Page 25: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Crystallography - Crystals

a

b

c

a

b

c

0 a 2a 3a 4a ta

b2b

c

2c

vc

ub

A crystal is a three dimensional stack of unit cells !

Page 26: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Crystallography - CrystalsDifferent unit cells:

a primitive unit cell

a unit cell centered in the planes

a body-centered unit cell

a face-centered unit cell

Page 27: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

DiffractionScattering by a crystal:

F0 (S) = fj (S) exp (2irj.S)

0 a 2a 3a 4a ta

b2b

c

2c

vc

ub

Page 28: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase
Page 29: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

DiffractionThe Result:

Page 30: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

DiffractionFourier Transformation

Page 31: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase
Page 32: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

-Stamp Collecting.

Page 33: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

-Crystallography of Large Complexes.-Time-Resolved Crystallography.-Locating Hydrogen Atoms.

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Page 35: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Substrate

Protein

Ligand

BINDING

REACTION

FUNCTION

STRUCTURAL CHANGE

Page 36: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Time Resolved Crystallography.

Page 37: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Mouse Prion Protein (PrPc)

NMR Structure

Page 38: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Protein Structure

Page 39: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase
Page 40: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase
Page 41: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

The Peptide Backbone Chain

Crosslinking - the disulphide bridge

Page 42: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Side-Chain Conformation

Page 43: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Linus Carl Pauling (1901-1994)

Nobel prize in Chemistry 1954 for his research into the nature of the chemical bond and its application to the elucidation of the structure of complex substances

Page 44: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

The Alpha Helix (Pauling & Corey, from alpha keratin)

Page 45: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase
Page 46: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Supersecondary Structure

Page 47: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Single helix:Alamethicin - a voltage-gated ion channel antibiotic

Tertiary Structure

Page 48: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Helix-turn Helix:Rop (RNA-Binding Protein) Four-Helix Bundle

Page 49: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Two Greek Keys (gamma crystallin)

Page 50: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

DNA-Binding Alpha Domains.

Page 51: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Cro repressor

Page 52: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

BETA - BARREL: PORIN

Page 53: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase
Page 54: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Alpha/Beta HORSHESHOEPlacental ribonuclease inhibitor: binds very strongly to any ribonuclease that leaks into the cytosol.

Page 55: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Same subunit found more than once:

Page 56: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Quaternary Structure:Aggregation of tertiary domains to form a quaternary structure.Photosynthetic reaction centre

Page 57: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Potassium Channel

Page 58: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Aquaporins

Page 59: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Strands from different protein chains associate to form 2o struct.

Page 60: End of Lecture 1 Diffraction Scattering – a system of two electrons: The two waves have equal amplitudes (because e 1 and e 2 are identical), but a phase

Same subunit associates